Genes Pathways Genomes

Kevin Munnelly, President and CEO, Gen9Stem Cells & Regenerative Medicine CongressSeptember 15, 2014

Synthetic Biology:A Perspective On Its Potential In Regenerative Medicine

Shameless Self Promotion

Gen9 Confidential2

20+ years in pharmaceuticals, life science tools and molecular diagnostics

Board Member

Fast, accurate, inexpensive gene sequencing using molecular motion

Providing functional water soluble variants of any GPCR or

membrane protein

Gen9 – Synthesizing the Future

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DrewEndy

Stanford

Bio-engineering

George Church

Harvard

Genetics

Joe Jacobson

MIT

Fabrication

Founded 200935 EmployeesLocated in Cambridge, MA, USA

Founders Investors

What is Synthetic Biology?

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"Synthetic biology is:a) the design and

construction of new biological parts, devices and systems and; b) the re-design of existing natural biological systems for useful purposes."

Synthetic Biology is Creating Future Factories…

…for the Production of Pharmaceuticals, Biofuels and Chemicals

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to Create Cellular Factories … to Make High Value ProductsUsing Synthetic Genes…

ChemicalsBio-Fuels EnzymesPharmaceuticals

Products of Synthetic Biology

Gen9 Confidential6

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Systems Biology• Construction and analysis of

synthetic regulatory networks• Sophisticated Perturbations

Bimolecular synthesis and fabrication• Optimized drug synthesis• Molecular-scale device fabrication

Biomedical• Artificial immune systems• Cancer and other disease therapies• Tissue generation and regulation• Biosensing• Diagnostics

Synthetic Biology in Medicine

The Progress of Synthetic Biology

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De novo Mycoplasma Genome synthesized

S. cerevisiae Chromosome synthesized

DNA

Prokaryote

Eukaryote

Tissue/Organ

Metazoan

Bi-stable Switch in E. coli

Designed Proteins

Synthetic CircuitsParts Libraries BioBricks repository established

2000

2005

2010

2015

2020

1995

What is Holding Us Back?

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Capacity in Base Pairs Per Year

3 Trillion bpAnnual

SequencingCapacity

300 Million bpAnnual

SynthesisCapacity

10,000X Discrepancy

The Opportunity: Reading vs. Writing DNA

Gene Synthesis: A new Demonstration of Moore’s Law

Disruptive Technologies

Integrated Circuit

Gen9 BioFab®

Simple Circuit Complex Circuit

Oligo Synthesis Gene Synthesis

Gene Synthesis Ecosystem

Gen9 Confidential12

Parts Library CAD

Bio-Informatics

BioFab®

On-Demand DNA Applications

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base

pairs 102 107105104103 106

single genes*

genetic circuits

Pathwaysgenome rewrite

minimal life

The Promise of Synthetic Biology

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Editing Genomes

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Advancements in technologies for precision editing

Guide RNACas9

Genomic DNA

Repair

Targeted Genome Editing

Synthetic Donor DNA

Creation of Pluripotent Cells

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Reprogramming:ERG, KLF4, SOX4, MYB, RORA,

HLF, HOXA5, HOXA9, and HOXA10

iPSFibroblast iPS Cells

Growth

Reprogramming:ERG, KLF4, SOX4, MYB, RORA,

HLF, HOXA5, HOXA9, and HOXA10

PLUS ADDITIONAL Factors

iPS+?Fibroblast

Growth

Pluripotent Reprogramming

Better Options for Therapy

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And Disease Treatment

Healthy DonorPatient

Engineered for:• Immune resistance?• Viral Resistance?• Additional disease

fighting capabilities

Engineered for:• Viral Resistance?• Additional disease

fighting capabilities• Anti Aging?

Recipient Patient

New Therapeutic Companies in Editing Space

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Need for Organs

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Each day, an average of 18 people die waiting for transplants due to organ shortage (>6500 year)

Data from optn.transplant.hrsa.gov and OPTN/SRTR Annual Report.** Data include deceased and living donors

Humanized Pigs?

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Immune Resistant Model Organisms

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Iterative genome editing approach to synthesize human compatible organs from animals

Target-cell isolation

Rapid toolbuilding

Accurateassessment

Targetingvalidation

Organ level functional

assays

Editing

Cas9

gRNA

Genome-wide gene discovery

Pig cloning and organ testing

Multipliable genome

engineering

Iterativehumanization of pig organs

DNA synthesis

CRISPR library

Engineered cell library

Can We Create Large Scale Functional Tissues?

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Defining, Writing and Manipulating Genetic Programs

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A Self-Timed Genetic Program for β cell Differentiation

From the Weiss lab (MIT Synthetic Biology Dept)

A Sampling of Cell Fate Regulators

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Targets for Synthetic Biology Reprogramming

J. Fujikura, et al., Genes and Development, Vol. 16,pp. 784‐789, 2002.R. Davis, H. Weintraub, A. B. Lassar, Cell Vol. 51, pp. 987‐1000, 1987.H. Niwa, et. al, Cell, Vol. 123(5), pp. 917‐929, 2005.Ericson, et al., Cell, Vol. 90, pp. 169‐180, 1997.G. Gu, et al., Development Vol. 129, pp. 2447‐2457, 2002.J. Wang, et al., Developmental Biology, Vol. 266, pp. 178‐189, 2004.N. Lavon, et al., Stem Cells, Vol. 24, pp. 1923‐1930, 2006.J. Heit, S. Karnik, S. Kim, Ann. Rev. Cell Dev. Biol. 22: 311‐338, 2006.D. Sinner, et al., Development, Vol. 131, pp. 3069‐3080, 2004J. Briscoe, et al., Nature, Vol. 398, p.622‐627, 1999.T. Jessell, Nature Reviews Genetics, Vol. 1, pp. 21‐29, 2000.M. McCormick, et al., Mol. and Cell. Bio., Vol. 16(10),pp. 5792‐5800, 1996M. Yasunaga, et al., Nature Biotech, Vol. 23(12), 1542‐1550, 2005.C. Vernochet, et al., FEBS Letters, Vol, 510, pp. 94‐98, 2002.

• Gata4, Gata6, Sox17

Endoderm

• MyoD

Muscle

• Cdx2

Trophectoderm

• Ngn3, Pax4, Nkx2.2, Pdx1

Pancreas

• Pparγ

Adipocytes

• Nkx2.2, Nkx6.1, Pax6, Ngn1

Neuronal

Path to Organ (re)Generation

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Reprogramming

Printed Organs

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Development Timeline for 3D Tissues/Organs

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Murphy and Atala. Nature Biotechnology Vol 32, No. 8 (2014)

Looking Ahead

• Advancement of developmental biology research

• iPS reprogramming

– Therapies

– Tissues

• Applications of amphibian based genetics?

• Generic and personalized model organisms

• SynBio + 3D printing

• Nuclei editing in Brain and neurons

• Anti Aging?

Gen9 Confidential29

Potential to unlock many of life’s mysteries!

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